Composition, synthesis, and use of new substituted pyran and pterin compounds

ABSTRACT

The present invention relates to substituted pterin compounds, their synthesis and use. In particular, the present invention relates to a new precursor compound and its analogs for synthesizing a new substituted pterin compound and its analogs. These new compounds are particularly suitable for treating molybdenum cofactor deficiency.

CROSS-RELATED APPLICATIONS

This patent application is a divisional of, and claims priority to,patent application Ser. No. 13/072,092 filed on Mar. 25, 2011, entitled“Composition, Synthesis, and Use of New Substituted Pyran and PterinCompounds” which issued as U.S. Pat. No. 8,378,123 on Feb. 19, 2013.

GOVERNMENT FUNDING

This invention was made with government support under grant#GM61555-04A1 awarded by the National Institutes of Health, NationalInstitute of General Medical Sciences. The government has certain rightsin the invention.

FIELD OF THE INVENTION

The present invention relates to new substituted pyran and pterincompounds, their synthesis and use. The new substituted pyran compoundscan be used as a precursor in the synthesis of the new substitutedpterin compounds. These new compounds are suitable for use in a widevariety of applications including, but not limited to, medicine andpharmaceuticals.

BACKGROUND OF THE INVENTION

Pterin is a heterocyclic compound composed of a pyrazine ring and apyrimidine ring (a pteridine ring system). The pyrimidine ring includesa carbonyl oxygen and an amino group. Pterins are derivatives of2-amino-4-oxopteridine with additional functional groups attached to thepyrazine ring. Pterins are known in the art and are used in a variousapplications. One of the most notable examples of pterin is folic acid.Another example is molybdopterin which is a substituted pteridine thatbinds molybdenum to give redox enzymes involved in biologicalhydroxylation and oxidation reactions. Pterins, substituted pterins, andderivatives thereof are of significant interest because of theirpotential uses in the fields of medicine and pharmaceuticals. It iscontemplated that pterins can be used as a means for therapy in treatingvarious medical conditions, such as cancer and molybdenum cofactordeficiency (MCD). Further, it is believed that pterins may possessantibacterial properties.

Pyran is a heterocyclic ring composed of five carbon atoms and oneoxygen atom. There are many pyran derivatives that are known in the artas important biological molecules.

Molybdenum is an essential trace element for virtually all life forms.It is central to a cofactor for a number of enzymes that catalyzeimportant chemical transformations. A biological form of molybdenumpresent in molybdenum-containing enzymes is known as the molybdenumcofactor. The molybdenum cofactor is a complicated molecule withmultiple redox-active components delicately balanced having thefollowing structure:

In some enzymes, the phosphate group substituent of the cofactor ismodified with a dinucleotide. The chemical synthesis of the molybdenumcofactor and its precursors is very challenging.

An absence of or deficiency in molybdenum cofactor (known as MCD) in thehuman body can lead to serious illness and death. MCD is a lethalautosomal recessive disease for which treatment and cure is a focus ofsignificant research efforts.

The biosynthetic pathway of molybdenum cofactor is a multi-step andevolutionarily conserved process involving four genes. Three of thesefour genes are linked to MCD. Patients are characterized by progressiveneurological damage, leading to early childhood death in most cases.Symptoms are mainly caused by the sulfite oxidase (SO) enzymedeficiency. The SO enzymes remove toxic sulfite in the human body toprotect the organs, particularly the brain, from an accumulation ofsulfite. A deficiency of SO enzymes causes excess sulfite to accumulatein plasma and serum, it crosses the blood-brain barrier, and rapidlytriggers neuronal death. Excess sulfite reacts with cystine, formingS-sulfocysteine, a potential agonist of glutamate receptors, which hasbeen implicated for observed seizures, convulsions, contractions andtwitching associated with MCD, causing damage of cortical neurons andloss of white matter.

There is a desire in the art to develop and synthesize new substitutedpyrans and pterins for use in various applications including medicineand pharmaceuticals, such as, but not limited to, the treatment of MCD.

SUMMARY OF THE INVENTION

Various aspects provide for compounds. Other aspects relate to thesynthesis of these compounds and their uses.

In one aspect, the present invention provides a compound comprising astructure:

wherein X is selected from the group consisting of oxygen and sulfur,and, R¹ and R² are each independently selected from the group consistingof hydrogen, substituted and non-substituted, aliphatic andnon-aliphatic hydrocarbon, branched and non-branched alkyl, cycloalkylhaving one or more rings, aryl including benzyl, phenyl, thienyl,indoyl, heteroaryl, phosphate including organic and inorganic,combinations thereof, and derivatives thereof.

In another aspect, the present invention provides a method for preparinga compound having a structure:

wherein X is selected from the group consisting of oxygen and sulfurand, R¹ and R² are each independently selected from the group consistingof hydrogen, substituted and non-substituted, aliphatic andnon-aliphatic hydrocarbon, branched and non-branched alkyl, cycloalkylhaving one or more rings, aryl including benzyl, phenyl, thienyl,indoyl, heteroaryl, phosphate including organic and inorganic,combinations thereof, and derivatives thereof. The method includesprotecting an alcohol group of an acetylene, deprotecting a hydrogen ofthe acetylene forming a corresponding acetylide compound, condensing theacetylide compound with an ester to form a substituted acetylene,combining the substituted acetylene with a sulfurating agent tointroduce a dithiolene group to form a dithiolene substituted furancompound, and treating the dithiolene substituted furan compound with anorganic compound or a mixture of organic compounds to form a substitutedpyran dithiolene compound.

In still another aspect, the present invention includes a compoundcomprising a structure selected from the group consisting of:

wherein X is selected from the group consisting of oxygen and sulfurand, R¹ and R² are each independently selected from the group consistingof hydrogen, substituted and non-substituted, aliphatic andnon-aliphatic hydrocarbon, branched and non-branched alkyl, cycloalkylhaving one or more rings, aryl including benzyl, phenyl, thienyl,indoyl, heteroaryl, phosphate including organic and inorganic,combinations thereof, and derivatives thereof.

In yet another aspect, the present invention provides a method ofpreparing a compound having a structure selected from the groupconsisting of:

wherein X is selected from the group consisting of oxygen and sulfurand, R¹ and R² are each independently selected from the group consistingof hydrogen, substituted and non-substituted, aliphatic andnon-aliphatic hydrocarbon, branched and non-branched alkyl, cycloalkylhaving one or more rings, aryl including benzyl, phenyl, thienyl,indoyl, heteroaryl, phosphate including organic and inorganic,combinations thereof, and derivatives thereof. The method includesstarting with a precursor compound having a structure:

wherein X is selected from the group consisting of oxygen and sulfurand, R¹ and R² are each independently selected from the group consistingof hydrogen, substituted and non-substituted, aliphatic andnon-aliphatic hydrocarbon, branched and non-branched alkyl, cycloalkylhaving one or more rings, aryl including benzyl, phenyl, thienyl,indoyl, heteroaryl, phosphate including organic and inorganic,combinations thereof, and derivatives thereof, treating the precursorcompound with o-phenylenediamine to form an open-ring compound,subjecting the open-ring compound to electrophilic activation leading toa corresponding ring closed quinoxaline derivative, and combining thequinoxaline derivative with pyrimidine in a condensation reaction toform the compound.

In another aspect, the present invention provides a therapeuticcomposition including the new substituted pterin compound of the presentinvention.

In yet another aspect, the present invention provides a method oftreating molybdenum cofactor deficiency including administering atherapeutically effective amount of the new substituted pterin compoundof the present invention.

BRIEF DESCRIPTION OF THE DRAWINGS

The various embodiments of the present disclosure may be betterunderstood when read with reference to the following figures.

FIG. 1 illustrates the synthesis of a substituted pyran-dithioleneprecursor compound.

FIG. 2 illustrates the synthesis of substituted quinoxaline and pterincompounds.

FIG. 3 illustrates ¹H NMR spectrum of compound 2 in CDCl₃.

FIG. 4 illustrates ¹³C NMR spectrum of compound 2 in CDCl₃.

FIG. 5 illustrates infrared spectrum (neat) of compound 2.

FIG. 6 illustrates infrared spectrum (neat) of4-phenyl-1,3-dithiolane-2-thione.

FIG. 7 illustrates ¹H NMR spectrum of compound 3 in CDCl₃.

FIG. 8 illustrates ¹³C NMR spectrum of compound 3 in CDCl₃.

FIG. 9 illustrates infrared spectrum (neat) of compound 5.

FIG. 10 illustrates high resolution mass spectrum (ESI⁺) of compound 3.

FIG. 11 illustrates ¹H NMR spectrum of compound 4 in CDCl₃.

FIG. 12 illustrates ¹³C NMR spectrum of compound 4 in CDCl₃.

FIG. 13 illustrates infrared spectrum (neat) of compound 4.

FIG. 14 illustrates high resolution mass spectrum (ESI⁺) of compound 4.

FIG. 15 illustrates ¹H NMR spectrum of compound 5 in CDCl₃.

FIG. 16 illustrates ¹³C NMR spectrum of compound 5 in CDCl₃.

FIG. 17 illustrates infrared spectrum (neat) of compound 5.

FIG. 18 illustrates high resolution mass spectrum (ESI⁺) of compound 5.

FIG. 19 illustrates ¹H NMR spectrum of compound 6 in CDCl₃.

FIG. 20 illustrates ¹³C NMR spectrum of compound 6 in CDCl₃.

FIG. 21 illustrates infrared spectrum (neat) of compound 6.

FIG. 22 illustrates high resolution mass spectrum (APCI⁺) of compound 6.

FIG. 23 illustrates ¹H NMR spectrum of compound 7 in CDCl₃.

FIG. 24 illustrates ¹³C NMR spectrum of compound 7 in CDCl₃.

FIG. 25 illustrates infrared spectrum (neat) of compound 7.

FIG. 26 illustrates high resolution mass spectrum (APCI⁺) of compound 7.

FIG. 27 illustrates ¹H NMR spectrum of compound 9 in CDCl₃.

FIG. 28 illustrates ¹³C NMR spectrum of compound 9 in CDCl₃.

FIG. 29 illustrates infrared spectrum (neat) of compound 9.

FIG. 30 illustrates high resolution mass spectrum (APCI⁺) of compound 9.

FIG. 31 illustrates ¹H NMR spectrum of compounds 14 and 15 in DMSO.

FIG. 32 illustrates ¹³C NMR spectrum of compounds 14 and 15 in DMSO.

FIG. 33 illustrates infrared spectrum (neat) of compounds 14 and 15.

FIG. 34 illustrates high resolution mass spectrum (ESI⁺) of compounds 14and 15.

FIG. 35 illustrates ¹H NMR spectrum of compound 8 in CDCl₃.

FIG. 36 illustrates ¹³C NMR spectrum of compound 8 in CDCl₃.

FIG. 37 illustrates infrared spectrum (neat) of compound 8.

FIG. 38 illustrates ¹H NMR spectrum of compounds 16 and 17 in DMSO.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to a new substituted pyran compound andits analogs that may be synthesized from readily available materials.This new compound can be used in various applications and in oneembodiment, is used as a precursor or starting material to synthesize anew substituted pterin compound and its analogs. The substituted pyrancompound includes a dithiolene group. The substituted pterin compoundincludes pyrimidine, pyrazine and dithiolene. The structures of thesecompounds are designed with the flexibility to have multiplesubstitution patterns. These compounds can have various uses and, inparticular, are suitable for the medical and pharmaceutical fields.

Other than the operating examples, or where otherwise indicated, allnumbers expressing quantities of ingredients, processing conditions andthe like used in the specification and claims are to be understood asbeing modified in all instances by the term “about”. Accordingly, unlessindicated to the contrary, the numerical parameters set forth in thefollowing specification and attached claims are approximations that mayvary depending upon the desired properties sought to be obtained. At thevery least, and not as an attempt to limit the application of thedoctrine of equivalents to the scope of the claims, each numericalparameter should at least be construed in light of the number ofreported significant digits and by applying ordinary roundingtechniques.

Notwithstanding that the numerical ranges and parameters setting forththe broad scope of the disclosure are approximations, the numericalvalues set forth in the specific examples are reported as precisely aspossible. Any numerical values, however, may contain certain errors,such as, for example, equipment and/or operator error, necessarilyresulting from the standard deviation found in their respective testingmeasurements.

Also, it should be understood that any numerical range recited herein isintended to include all sub-ranges subsumed therein. For example, arange of “1 to 10” is intended to include all sub-ranges between (andincluding) the recited minimum value of 1 and the recited maximum valueof 10, that is, having a minimum value equal to or greater than 1 and amaximum value of less than or equal to 10.

The present disclosure describes several different features and aspectsof the invention with reference to various exemplary non-limitingembodiments. It is understood, however, that the invention embracesnumerous alternative embodiments, which may be accomplished by combiningany of the different features, aspects, and embodiments described hereinin any combination that one of ordinary skill in the art would finduseful.

In one aspect, the present disclosure relates to the development of anew substituted pyran compound having a structure including at least twofused rings. One ring includes a five-membered ring (e.g., protecteddithiolene) and another ring includes a six-membered ring (e.g., pyran).The general structure of the new substituted pyran compound isrepresented by structure I.

In structure I, X is selected from oxygen (O) and sulfur (S). Further,in structure I, R¹ and R² are each independently selected from hydrogen,substituted and non-substituted, aliphatic and non-aliphatichydrocarbon, branched and non-branched alkyl, such as C₁-C₆ alkyl,hydroxy C₁-C₆ alkyl, alkenyl, alkynyl, cycloalkyl having one or morerings, aryl including benzyl, phenyl, thienyl, indoyl, heteroaryl,phosphate including organic and inorganic, combinations thereof, andderivatives thereof R¹ and R² may be the same or different. As usedherein, the term “aryl” includes an aromatic ring (i.e., a singlearomatic ring) or a ring system (i.e., a polycyclic aromatic ringsystem) in which all ring atoms are carbon. In one embodiment, R¹ and R²are each independently selected from methyl (CH₃), hydroxyl ethyl,organic phosphate, and inorganic phosphate. In another embodiment, R¹and R² are both methyl or hydroxy ethyl or phosphate.

According to various embodiments, the substituted pyran having thegeneral structure of structure I, may be readily synthesized usingorganic chemistry techniques. For example, the synthesis of variousembodiments of the substituted pyran is described herein. It should benoted that the featured embodiments are intended to be exemplary and arein no way limiting to the scope of the precursor as described herein.The synthetic approach for the substituted pyran includes beginning withan acetylene compound. The acetylene compound can be selected from thosethat are known in the art and commercially available. The acetylenecompound is soluble in an organic solvent. The organic solvent can beselected from those that are known in the art and commerciallyavailable. In one embodiment, the organic solvent is dichloromethane. Inanother embodiment, the acetylene compound has the general structurerepresented by structure II.

In structure II, R¹ and R² are each independently selected fromhydrogen, substituted and non-substituted, aliphatic and non-aliphatichydrocarbon, branched and non-branched alkyl, such as C₁-C₆ alkyl,hydroxy C₁-C₆ alkyl, alkenyl, alkynyl, cycloalkyl having one or morerings, aryl including benzyl, phenyl, thienyl, indoyl, heteroaryl,phosphate including organic and inorganic, combinations thereof, andderivatives thereof R¹ and R² may be the same or different. As usedherein, the term “aryl” includes an aromatic ring (i.e., a singlearomatic ring) or a ring system (i.e., a polycyclic aromatic ringsystem) in which all ring atoms are carbon. In one embodiment, R¹ and R²are each independently selected from methyl (CH₃), hydroxyl ethyl,organic phosphate, and inorganic phosphate. In another embodiment, R¹and R² are both methyl or hydroxy ethyl or phosphate.

Further, the synthetic approach includes protecting the hydroxyl group(e.g., alcohol) of the acetylene compound (as shown in structure II). Inone embodiment, an organic compound, such as dihydropyran, is combinedwith the acetylene compound. The combination of these compounds can becarried out in the presence of catalyst. The catalyst can be selectedfrom those that are known in the art and commercially available. In oneembodiment, p-toluensulfonic acid is used in a catalytic amount. Theresulting compound is deprotonated with an organic reagent and condensedwith an ester. The organic reagent and the ester can be selected fromthose that are known in the art and commercially available. In oneembodiment, the protected acetylene compound is combined with an organicsolvent, such as diethyl ether. To this mixture n-butyl lithium is addedand ethyl 2,2-diethoxyacetate is subsequently added to form asubstituted acetylene compound. A dithiolene unit is then introduced,for example, by adding a sulfurating agent, and a dithiolene substitutedcompound having two, five-member rings is formed. The acetal group onthe furan ring is deprotected ultimately leading to a substitutedsix-member pyran ring.

A specific example is discussed in detail in FIG. 1. As illustrated inFIG. 1, the synthetic approach begins with the commercially availableacetylene, 2-methyl-3-butyn-2-ol 1 treated with tetrahydropyran (THP) toprotect the hydroxyl group (e.g., alcohol). The acetylene 3-methyl-3tetrahydropyranyloxy-butyne2-(2-methylbut-3-yn-2-yloxy)tetrahydro-2H-pyran 2 is formed and is thendeprotonated with n-butyl lithium and condensed with the ester ethyl2,2-diethoxyacetate to form the substituted acetylide1,1-diethoxy-5-methyl-5-(tetrahydro-2H-pyran-2-yloxy)hex-3-yn-2-one 3. Adithiolene unit is then introduced by sulfurating the substitutedacetylene 3 with the sulfurating agent 4-phenyl-1,3-dithiolane-2-thioneto produce the dithiolene substituted compound4-(diethoxymethyl)-4-hydroxy-6,6-dimethyl-4,6-dihydro-[1,3]dithiolo[4,5-c]furan-2-thione4 having a five-member dihydrofuran ring and acetal functionality. Thecompound 4 is treated with lutidine and trimethylsilyl trifluoromethanesulfonate (TMSOTf) to deprotect the acetal and to form the six-memberring of the compound6-hydroxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,5-c]pyran-7(6H)-one 6and 6-hydroxy-4,4-dimethyl-4H-[1,3]dithiolo[4,5-c]pyran-2,7(6H)-dione 8.

NMR and IR spectroscopies and HRMS can be used to determine thestructure of compound 4 consistent with the presence of the dithioleneunit, the five-membered dihydrofuran ring and the acetal functionality.Without intending to be bound by any particular theory, it is believedthat the presence of the keto-aldehyde functionality can make thecompound particularly reactive towards condensation reactions.

Each of the compounds 6 and 8 can be used as a precursor in thesynthesis of a substituted pterin compound. The substituted pterincompound can have the general structure represented by structures IIIaand IIIb.

In structures IIIa and IIIb, X is selected from oxygen or sulfur and, R¹and R² are each independently selected from hydrogen, substituted andnon-substituted, aliphatic and non-aliphatic hydrocarbon, branched andnon-branched alkyl, such as C₁-C₆ alkyl, hydroxy C₁-C₆ alkyl, alkenyl,alkynyl, cycloalkyl having one or more rings, aryl including benzyl,phenyl, thienyl, indoyl, heteroaryl, phosphate including organic andinorganic, combinations thereof, and derivatives thereof R¹ and R² maybe the same or different. As used herein, the term “aryl” includes anaromatic ring (i.e., a single aromatic ring) or a ring system (i.e., apolycyclic aromatic ring system) in which all ring atoms are carbon. Inone embodiment, R¹ and R² are each independently selected from methyl(CH₃), hydroxyl ethyl, organic phosphate, and inorganic phosphate. In apreferred embodiment, R¹ and R² are both methyl or hydroxy ethyl orphosphate.

A specific example is discussed in detail in FIG. 2. As illustrated inFIG. 2, the synthetic approach is started with the precursor compound 6or 8. For example, it is shown in FIG. 2 that the precursor compound 6can be combined with an organic solvent, such as dichloromethane. Thecompound 6 in solution can be reacted with o-phenylenediamine to formthe open-ring compound4-(2-hydroxypropan-2-yl)-5-(quinoxalin-2-yl)-1,3-dithiole-2-thione 9.Subsequently, closed-ring compounds 10 and 11 (e.g., pyran) can beformed through electrophilic activation with benzyl chloroformate(Cbz-Cl). The compound 11 is a fully oxidized fluorescent quinoxalinederivative. The compound 10 is a reduced quinoxaline with the nitrogenprotected with a carboxybenzyl group.

Condensation reactions can be conducted using pyrimidines. Thepyrmidines can include protected pyrimidines, such as protectedpyrimidine diamine, and unprotected pyrimidines, such as unprotectedpyrimidine diamines. In FIG. 2, the precursor compound 6 can becondensed using a partially protected pyrimidine to faun the open-ringisomer compounds 12 and 13. Further, in FIG. 2, the precursor compounds6 and 8 can be condensed using the unprotected pyrimidine4-hydroxy-2,5,6-triaminopyrimidine sulfate to form the open-ring isomercompounds 14 and 15, and, 16 and 17, respectively. The use ofpyrimidines can result in the reaction being completed in a relativelyshort period of time and yielding significant rates of conversion. Inone embodiment, the reaction can be completed in approximately 30minutes.

It is believed that isomer compounds 14, 15, 16 and 17 which are in aring-open form can be converted to a ring-closed form. In oneembodiment, the synthetic approach may include reacting compound 14 or15 or 16 or 17 with a protecting agent to protect the nitrogen atoms andcyclize the pyran ring. This can be reduced by a reducing agent tofurnish a reduced pterin.

The new substituted pyran and pterin compounds can have various uses andcan be suitable for use the fields of medicine, pharmaceuticals, andrelated fields. It is contemplated that these compounds can be used astherapy for a wide range of medical conditions, such as, but not limitedto, cancer, vitamin deficiency, such as, folic acid deficiency, mineraldeficiency and molybdenum cofactor deficiency. It is furthercontemplated that the compounds of the present invention can beadministered to a human in therapeutic doses to treat these medicalconditions. In one embodiment, the substituted pterin compound ispresent in a composition that is administered in a therapeutic amount toa patient having a medical condition as described above. Furthermore, itis believed that the new substituted pterin compounds may exhibitantibacterial properties and therefore, may be used as an antibacterialagent.

As used herein, the term “therapeutically effective amount” refers tothat amount of any of the new substituted pterin compounds of thepresent invention incorporated in a pharmaceutical composition which isrequired to bring about a desired effect. As will be understood by oneskilled in the art, a therapeutically effective amount of the newsubstituted pterin compounds can be administered by any means known inthe art including, but not limited to, injection, parenterally, orally,or where appropriate, topically.

EXAMPLES Materials and Instrumentation

The chemicals used in the following examples were purchased from AldrichChemical Company or ACROS Chemical Company and were used without furtherpurification. Ether and dichloromethane were dried using solventpurification system and other solvents were used as received. The radialchromatography was performed with a Chromatotron using silica gel (5-40μm) purchased from EMD Chemicals. The ¹H and ¹³C NMR spectra wererecorded on Bruker Avance 400 and Bruker Avance 500 spectrometersoperating at 400 MHz and 500 MHz. The IR spectra were recorded using aNicolet 380 FT spectrophotometer. High resolution mass spectra wererecorded on an Agilent 6200 time of flight LC MS system using a nano ESIand APCI-TOF interface.

Synthesis of 3-methyl-3tetrahydropyranyloxy-butyne2-(2-methylbut-3-yn-2-yloxy)tetrahydro-2H-pyran (Compound 2)

To a cooled (−20° C.) solution of 2-methyl-3-butyn-2-ol 1 (15 g, 0.17moles) in 150 mL of dry CH₂Cl₂, 24.3 g (0.28 mole) dihydropyran wasadded and a catalytic amount of p-toluensulfonic acid (a few crystals).The progress of the reaction was monitored by thin layer chromatography(TLC) using silica gel as the stationary phase and CH₂Cl₂ as the mobilephase, and the spots were detected by concentrated H₂SO₄ treatment. Theresulting solution was stirred for 80 minutes and washed with asaturated solution of NaHCO₃ (3×200 mL). The organic layer was separatedand dried over anhydrous MgSO₄. The solvent was removed at a reducedpressure and the resulting oil was purified by vacuum distillation toyield the target compound 2. Yield: 27.2 g (0.16 mol; 91%). ¹H-NMRspectrum in CDCl₃ (δ, ppm): 5.06 (m, 1H, THP), 3.95 (m, 2H, THP), 3.50(m, 2H, THP), 2.43 (s, 1H, alkyne) 1.85 (m, 2H, THP), 1.70 (m, 2H, THP),1.51 (s, 6H, Me). ¹³C NMR spectrum in CDCl₃ (δ, ppm): 95.97, 86.25,86.24, 71.78, 70.72, 63.12, 31.80, 30.47, 29.68, 25.29, 20.29. IRspectra (neat, cm⁻¹): 3295, 2941, 2108, 1466, 1380.

Synthesis of 4-phenyl-1,3-dithiolane-2-thione

To a solution of 21.2 g KOH, dissolved in 160 mL MeOH was added 21 mL(0.33 mmol) CS₂, at −20° C. To this solution, 15 mL (0.12 mmol) styreneoxide was added slowly and the reaction mixture was stirred for 6 hoursat 0° C. The target compound, 4-phenyl-1,3-dithiolane-2-thione,precipitated as a yellow solid, which was filtered and washed withmethanol and water. The solid was dissolved in 50 mL of CH₂Cl₂, driedwith anhydrous MgSO₄, filtered, and the organic solvent was removedunder reduced pressure to yield a yellow solid. Yield: 13.8 g (0.06mmol, 52%). IR spectra (neat, cm⁻¹): 3027, 1487, 1442, 1045, 878, 690.

Synthesis of1,1-diethoxy-5-methyl-5-(tetrahydro-2H-pyran-2-yloxy)hex-3-yn-2-one(Compound 3)

In a 250 mL round bottom flask, 9.5 mL of n-butyl lithium (2.5 Msolution in hexane, 23.8 mmol) was added via a syringe to a pre-cooledsolution (−78° C.) of 2-(2-methylbut-3-yn-2-yloxy)-tetrahydro-2H-pyran 2(4.0 g, 23.8 mmol) in 100 mL of Et₂O. The resulting solution was stirredfor 30 min at −78° C., then ethyl 2,2-diethoxyacetate (4.9 g, 27.7 mmol)was added with a syringe, and the stirring continued for 12 hours. Thereaction mixture was poured in an ice-cold saturated aqueous solution ofNH₄Cl. The aqueous layer was extracted with Et₂O (3×50 mL). The organicsolvent was separated with a separating funnel and dried with anhydrousMgSO₄. The solvent was removed under reduced pressure and the resultingpale yellow oil was purified by radial chromatography (stationary phasesilica gel, hexane/AcOEt 90:10) to provide1,1-diethoxy-5-methyl-5-(tetrahydro-2H-pyran-2-yloxy)hex-3-yn-2-one 3 asa colorless liquid. Yield: 3.2 g (10.7 mmol; 45%). ¹H-NMR in CDCl₃ (δ,ppm): 5.05 (1H, q, J=5.0, 3.5 Hz, THP), 4.73 (1H, s, acetal), 3.92 (1H,m, THP), 3.69 (2H, m, CH₂, acetal), 3.62 (2H, m, CH₂, acetal), 3.50 (1H,m, THP), 1.82 (1H, m, THP), 1.71 (1H, m, THP), 1.58 (3H, s, CH₃), 1.54(3H, s, CH₃), 1.52 (4H, m, THP), 1.25 (6H, t, J=7.0 Hz, CH₃, acetal).¹³C-NMR in CDCl₃ (δ, ppm): 182.6, 101.1, 97.87, 96.10, 81.25, 70.57,62.95, 62.64, 62.61, 31.47, 29.45, 28.85, 25.06, 19.92, 14.90 IR (neat,cm⁻¹): 2978, 2937, 2872, 2214, 1687, 1074. HR ESIMS⁺ with acetonitrileas the mobile phase, (m/z): 299.1837 (M⁺, C₁₆H₂₆O₅, 299.1853), 321.1660(m/z) ([M+Na]⁺, C₁₆H₂₆O₅Na, 321.1672).

Synthesis of4-(diethoxymethyl)-4-hydroxy-6,6-dimethyl-4,6-dihydro-[1,3]dithiolo[4,5-c]furan-2-thioneand2,2-diethoxy-1-(5-(2-(tetrahydro-2H-pyran-2-yloxy)propan-2-yl)-2-thioxo-1,3-dithiol-4-yl)ethanone(Compounds 4 and 5, Respectively)

A mixture of1,1-diethoxy-5-methyl-5-(tetrahydro-2H-pyran-2-yloxy)hex-3-yn-2-one 3(1.5 g, 5.0 mmol) and 4-phenyl-1,3-dithiolane-thione (5.3 g, 25.0 mmol)was heated under N₂ at 130° C. for 1 hour. The reaction was monitored bythin layer chromatography using silica gel as the stationary phase anddichloromethane as the mobile phase. The color of the reaction mixturechanged from yellow to dark orange. After completion of the reaction, asevidenced by TLC, the reaction was stopped, cooled, and the targetcompound 4 was purified by radial chromatography using silica gel as thestationary phase. The target compound 4 eluted as the third fractionwith a mixture of dichloromethane and hexanes (60:40). The first bandwas the excess 4-phenyl-1,3-dithiolane-thione, and the second bandyielded2,2-diethoxy-1-(5-(2-(tetrahydro-2H-pyran-2-yloxy)propan-2-yl)-2-thioxo-1,3-dithiol-4-yl)ethanone5 which was further converted to4-(diethoxymethyl)-4-hydroxy-6,6-dimethyl-4,6-dihydro-[1,3]dithiolo[4,5-c]furan-2-thione4.

2,2-diethoxy-1-(5-(2-(tetrahydro-2H-pyran-2-yloxy)propan-2-yl)-2-thioxo-1,3-dithiol-4-yl)ethanone5 was isolated as a yellow oil. Yield: 80 mg (0.2 mmol, 4%). ¹H-NMRspectrum in CDCl₃ (δ, ppm): 4.96 (1H, dd; J, 3.7, 4.7 Hz, THP), 4.81(1H, s, acetal), 3.91 (1H, m, THP), 3.75 (2H, m, CH₂, acetal), 3.61 (2H,m, CH₂, acetal), 3.50 (1H, m, THP), 1.83 (1H, m, THP), 1.71 (3H, s,CH₃), 1.67 (3H, s, CH₃), 1.56 (5H, m, THP), 1.23 (6H, dt, J=0.6, 7.0,CH₃, acetal). ¹³C NMR in CDCl₃ (δ, ppm): 211.91, 186.77, 166.07, 130.57,102.31, 93.94, 80.90, 63.98, 63.88, 62.49, 31.27, 27.38, 25.26, 24.70,19.49, 15.10, 15.08. IR (neat, cm⁻¹): 2974, 2937, 2872, 1703, 1053. HRESIMS⁺ (m/z): 407.1005 (M⁺, C₁₇H₂₆O₅S₃, 407.1015).

The third band yielded4-(diethoxymethyl)-4-hydroxy-6,6-dimethyl-4,6-dihydro-[1,3]dithiolo[4,5-c]furan-2-thione4 as a dark yellow liquid which solidified after sometime. Yield: 0.642g (2.0 mmol, 40%). ¹H-NMR in CDCl₃ (δ, ppm): 4.44 (1H, s, acetal), 3.97(1H, s, —OH), 3.81 (3H, m, acetal), 3.59 (1H, m, CH₂, acetal), 1.62 (3H,s, CH₃), 1.52 (3H, s, CH₃), 1.28 (3H, t; J, 7.0 Hz, CH₃, acetal), 1.24(3H, t; J, 7.0 Hz, CH₃, acetal). ¹³C NMR in CDCl₃ (δ, ppm): 219.08,150.68, 136.55, 106.34, 103.56, 87.11, 65.87, 65.21, 29.64, 28.53,15.29, 14.99. IR (neat, cm⁻¹): 3419, 2974, 2884, 1057. HR ESIMS⁺ (m/z):323.0452 (M⁺, C₁₂H₁₈O₄S₃, 323.0440).

Synthesis of4-(diethoxymethyl)-4-hydroxy-6,6-dimethyl-4,6-dihydro-[1,3]dithiolo[4,5-c]furan-2-thione(Compound 4)

10 mg (0.024 mmol) of2,2-diethoxy-1-(5-(2-(tetrahydro-2H-pyran-2-yloxy)propan-2-yl)-2-thioxo-1,3-dithiol-4-yl)ethanone5 was dissolved in 5 mL of CH₂Cl₂ containing 0.1 mL CF₃COOH. Thereaction mixture was stirred for 12 hours, washed with 20 mL ofsaturated solution of NaHCO₃ and, the organic layer was separated anddried in vacuum to yield the target compound 4 as a solid material.Yield: 7.2 mg (0.02 mmol, 90%).

Synthesis of6-hydroxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,5-c]pyran-7(6H)-oneand6-ethoxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,4-c]pyran-7(6H)-one(Compounds 6 and 7, Respectively)

245 mg (0.76 mmol) of4-(diethoxymethyl)-4-hydroxy-6,6-dimethyl-4,6-dihydro-[1,3]dithiolo[4,5-c]furan-2-thione4 was dissolved in 2.5 mL of dry CH₂Cl₂ under nitrogen atmosphere insidea Nexus One inert atmosphere box. To this solution, 758 μl (6.57 mmol)of 2,6-lutidine was added followed by addition of 843 μl (4.65 mmol) oftrimethylsilyl trifluoromethanesulfonate. The mixture was stirred for 17hours at room temperature and 3 mL water, 3 mL acetone, and 1 dropformic acid were added sequentially. The mixture was refluxed for 1hour. The organic material was extracted with CH₂Cl₂ and washed with asaturated solution of NaHCO₃. The organic layer was separated and driedover anhydrous MgSO₄. The solvent was removed by vacuum and theresulting solid was purified by radial chromatography using CH₂Cl₂ asthe mobile phase. The first yellow fraction was the6-ethoxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,4-c]pyran-7(6H)-one 7isolated as a yellow oil. Yield: 12 mg (0.05 mmol, 6%). ¹H NMR in CDCl₃(δ, ppm): 5.05 (1H, s, acetal), 3.95 (1H, m, CH₂, acetal), 3.68 (m, 1H,CH₂, acetal), 1.78 (3H, s, CH₃), 1.60 (3H, s, CH₃), 1.26 (3H, t, J=7.1Hz, CH₃, acetal). ¹³C NMR in CDCl₃ (δ, ppm): 208.78, 179.10, 167.34,133.35, 98.23, 75.50, 65.57, 31.90, 31.23, 15.24. IR (neat, cm⁻¹): 2978,2925, 2888, 2691, 1041. HR APCIMS⁺ (m/z): 277.0012 (M⁺, C₁₀H₁₂O₃S₃,277.0021).

The second fraction was6-hydroxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,5-c]pyran-7(6H)-one 6isolated as a dark yellow liquid that solidified. Yield: 146.3 mg (0.59mmol, 78%). ¹H-NMR in CDCl₃ (δ, ppm): 5.50 (1H, d; J, 6.0 Hzhemiacetal), 3.54 (1H, d; J, 6.0 Hz, OH), 1.75 (3H, s, CH₃), 1.69 (3H,s, CH₃). ¹³C NMR in CDCl₃ (δ, ppm): 207.93, 181.1, 167.6, 132.31, 91.28,75.71, 31.37, 28.51. IR (neat, cm⁻¹): 3366, 2978, 2929, 2869, 1679,1078, 1017. HR APCIMS⁺ (m/z): 248.9700 (M⁺, C₈H₈O₃S₃, 248.9708).

Synthesis of6-hydroxy-4,4-dimethyl-4H-[1,3]dithiolo[4,5-c]pyran-2,7(6H)-dione(Compound 8)

13 mg (0.05 mmol) of6-hydroxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,5-c]pyran-7(6H)-one 6was dissolved in acetone (5 mL) and added to a solution of Hg(OAc)₂ (50mg, 0.15 mmol) in 1 ml of acetic acid. The color of the reaction mixturechanged very fast from yellow to white. The solution was stirred for 10minutes and then 20 mL of water was added. The resulting mixture wasextracted with CH₂Cl₂ (3×5 mL). The organic layer was separated and thenwas washed with saturated aqueous NaHCO₃ (10 mL). The organic layer wasseparated again and the aqueous layer was re-extracted again with CH₂Cl₂(2×5 mL). The organic layers were combined and evaporated to get6-hydroxy-4,4-dimethyl-4H[1,3]dithiolo[4,5-c]pyran-2,7(6H)-dione 8 as apale beige oil. Yield: 7 mg (0.03 mmol, 57%). ¹H NMR in CDCl₃ (δ, ppm):5.51 (1H, s, hemiacetal), 1.74 (3H, s, CH₃), 1.68 (3H, s, CH₃). ¹³C NMRin CDCl₃ (δ, ppm): 187.09, 181.81, 159.93, 124.38, 91.15, 76.16, 31.47,28.27. IR (neat, cm⁻¹): 3419, 2978, 2929, 2864, 1670, 1625, 1548.

Synthesis of4-(2-hydroxypropan-2-yl)-5-(quinoxalin-2-yl)-1,3-dithiole-2-thione(Compound 9)

To a solution of6-hydroxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,5-c]pyran-7(6H)-one 6(101.0 mg, 0.41 mmol) in CH₂Cl₂ (15 mL), 136.6 mg (1.26 mmol) ofo-phenylenediamine was added, the resulting solution was refluxed for 1hour. The reaction mixture was cooled to room temperature and thesolvent was evaporated under reduced pressure. The residue was purifiedby radial chromatography using CH₂Cl₂ as a mobile phase that yielded4-(2-hydroxypropan-2-yl)-5-(quinoxalin-2-yl)-1,3-dithiole-2-thione 9 asa yellow-orange solid (119.6 mg, 0.37 mmol, 92.0%). ¹H NMR in CDCl₃ (δ,ppm): 9.01 (1H, s), 8.18 (1H, m, ArH), 8.07 (1H, m, ArH), 7.89 (2H, m,ArH), 1.59 (6H, s, CH₃); ¹³C NMR in CDCl₃ (δ, ppm): 207.8, 158.0, 145.1,144.3, 141.9, 140.0, 133.81, 132.3, 131.9, 129.8, 128.7, 72.3, 31.0; IR(neat, cm⁻¹): 3199, 3007, 2962, 2917, 1531, 1045, 759, 571, 457; HRAPCIMS⁺ (m/z): 321.0179 (M⁺, C₁₄H₁₂N₂OS₃, 321.0185).

Formation of the Pyran Ring in Compound 9

To compound 9 (34.6 mg, 0.11 mmol in 2 ml dichloromethane), 8 ml ofbenzylchloroformate (56 mmol) was added. The reaction mixture wasstirred at room temperature for a short time, the solution wasevaporated to dryness in vacuum in a span of 48 h. The residue waspurified by preparative TLC (silica gel, stationary phase;dichloromethane, mobile phase). The first fraction was collected to becompound 10 in ˜16% yield, and second band yielded compound 11 in 38%yield. Characterization data for compound 10, ¹H-NMR in CDCl₃ (δ, ppm):8.17 (1H, dd; J, 1.2, 8.5 Hz, ArH), 7.45 (1H, dd; J, 1.6, 7.5 Hz, ArH),7.38 (m, ArH), 7.13 (1H, td; J, 1.4, 7.78 Hz, ArH), 6.16 (1H, s; H-5a),5.32 (2H, d; J, 2.8 Hz, COHCHPh), 1.48 (3H, s., CH₃), 1.43 (3H, s, CH₃).Characterization data for compound 11, ¹H-NMR s in CDCl₃ (δ, ppm): 7.96(1H, d; J, 1.4, 8.0 Hz), 7.83 (1H, d; J, 1.4, 8.0 Hz), 7.68 (1H, td; J,8.0, 1.4 Hz), 7.61 (1H, td; J, 8.0, 1.4 Hz), 1.84 (6H, s, CH₃). ¹³C-NMRin CDCl₃ (δ, ppm): 209.7; 152.6; 149.4; 141.2; 139.6; 133.3; 133.1;130.8; 128.7; 128.2; 127.5; 80.9; 30.1. IR (neat, cm⁻¹): 1573, 1492,1463, 1409, 1365. APCIMS⁺ (m/z): 318.85 (C₁₄H₁₁N₂OS₃, 319.00). UV-vis inacetonitrile (λ_(max), nm (ε, M⁻¹ cm⁻¹)): 227 (17785), 257 (12623),391(16664), 410 (20682).

Synthesis of a Pterin Dithiolene Compounds,7-(5-(2-hydroxypropan-2-yl)-2-thioxo-1,3-dithiol-4-yl)-2-morpholinopteridin-4(3H)-oneand7-(5-(2-hydroxypropan-2-yl)-2-thioxo-1,3-dithiol-4-yl)-2-morpholinopteridin-4(3H)-one(Compounds 12 and 13, Respectively)

4.5 mg of compound 6 (0.018 mmol) dissolved in DMSO was reacted withmorpholine protected 5,6 diaminopyrimidine in a NMR tube for 40 minsmaintaining the temperature in a boiling water bath. The reactionmixture was followed by NMR spectroscopy, transferred into a flaskcontaining a small amount of water. It was then extracted with a smallamount of dichloromethane, the organic solvent was removed in vacuum.Yield, 5.5 mg (0.013 mmol, 72%) of compounds 12 and 13. ¹H-NMR in DMSO(δ, ppm): 12, 8.92(1H, s, H—C7), 1.41 (6H, s, CH₃); 13, 8.66 (1H, s,H—C6), 1.43 (6H, s, CH₃). HR ESI MS⁺ (m/z): 424.0581 (M⁺, C₁₆H₁₉N₅O₃S₃,424.0566).

Synthesis of2-amino-6-(5-(2-hydroxypropan-2-yl)-2-thioxo-1,3-dithiol-4-yl)pteridin-4(3H)-oneand2-amino-7-(5-(2-hydroxypropan-2-yl)-2-thioxo-1,3-dithiol-4-yl)pteridin-4(3H)-one(Compounds 14 and 15, Respectively)

45.0 mg 4-hydroxy-2,5,6-triaminopyrimidine sulfate (0.20 mmol) wassuspended in 5 mL of water (pH, 2.3) and solid sodium bicarbonate wasadded slowly to adjust the pH of the solution to 4.5. To this solution,solid sodium bisulfite was added slowly and the pH was adjusted to 5.9.To this solution, a solution of6-hydroxy-4,4-dimethyl-2-thioxo-4H-[1,3]dithiolo[4,5-c]pyran-7(6H)-one 6(50 mg, 0.20 mmol, in 5 mL ethanol) was added drop wise, and thereaction mixture was stirred at room temperature for 25 hours. Theresulting slurry was centrifuged. The solid was washed with waterfollowed by ethanol, and dried in vacuum to give yellow colored finepowder. The ratio of the two isomers, 14:15 was ˜82:18, as evidenced by¹H NMR spectrum. Yield: 30 mg (0.085 mmol, 42%). ¹H-NMR in DMSO (δ,ppm): 14, 8.89 (1H, s, H—C7), 1.44 (6H, s, CH₃); 15, 8.60 (1H, s, H—C6),1.46 (6H, s, CH₃); ¹³C NMR in DMSO (δ, ppm): 211.45, 161.60, 157.82,157.62, 155.89, 151.32, 138.82, 133.71, 128.91, 73.75, 31.9. IR (neat,cm⁻¹): 3346, 3364, 3129, 3966, 2921, 2827, 1699, 1536, 1045, 624, 530,440. HR ESI MS⁺ (m/z): 354.0122 (M⁺, C₁₂H₁₁N₅O₂S₃, 354.0148).

Synthesis of2-amino-6-(5-(2-hydroxypropan-2-yl)-2-oxo-1,3-dithiol-4-yl)pteridin-4(3H)-oneand2-amino-7-(5-(2-hydroxypropan-2-yl)-2-oxo-1,3-dithiol-4-yl)pteridin-4(3H)-one(Compounds 16 and 17, Respectively)

A small amount of6-hydroxy-4,4-dimethyl-4H-[1,3]dithiolo[4,5-c]pyran-2,7(6H)-dione, 8 wasdissolved in DMSO-d₆. Then, 4-hydroxy-2,5,6-triaminopyrimidine sulfateand solid sodium bisulfite were suspended in this solution. The color ofthe reaction mixture changed from colorless to green and then yellow.The mixture was heated on a water bath for 15 min. The mixture wasanalyzed by NMR. ¹H-NMR in DMSO (δ, ppm): 16, 8.84 (1H, s, H—C7), 1.39(6H, s, CH₃); 17, 8.55 (1H, s, H—C6), 1.43 (6H, s, CH₃).

It will be appreciated by those skilled in the art that changes could bemade to the embodiments described above without departing from the broadinventive concept thereof. It is understood, therefore, that thisinvention is not limited to the particular embodiments disclosed, but itis intended to cover modifications that are within the spirit and scopeof the invention, as defined by the appended claims.

What is claimed is:
 1. A method for preparing a compound of structure I:

wherein X is selected from the group consisting of oxygen and sulfurand, R¹ and R² are each independently unsubstituted C₁-C₆ alkyl,comprising: a. obtaining an acetylene compound of structure II:

wherein R¹ and R² are defined as above for structure I; b. combining thecompound of structure II with a hydroxyl protecting group which forms acompound of structure III:

wherein R¹ and R² are defined as above for structure I, and Y is ahydroxyl protecting group; c. deprotonating the compound of structureIII and condensing the compound of the structure III with ethyl2,2-diethoxy acetate ester which forms a substituted acetylene compoundof structure IV:

wherein R¹ and R² are defined as above for structure I, Et is ethyl, andY is as defined above for structure III d. combining the substitutedacetylene compound of structure IV with sulfurating agent4-phenyl-1,3-dithiolane-2-thione to form a dithiolene substitutedcompound of structure V:

wherein R¹ and R² are defined as above for structure I, and Et is asdefined above for structure IV; and e. deprotecting acetal in thecompound of structure V to form the compound of structure I.
 2. Themethod of claim 1 wherein the acetylene compound is2-methyl-3-butyn-2-ol.
 3. The method of claim 1 wherein step b furthercomprises catalyst.
 4. The method of claim 3 wherein the catalystcomprises p-toluensulfonic acid.
 5. The method of claim 1 wherein thehydroxyl protecting group in steps b and c is tetrahydropyran.
 6. Themethod of claim 1 wherein the deprotecting of the acetal in the compoundof structure V comprises combining the compound of structure V withtrimethylsilyl trifluoromethane sulfonate.